A membrane-associated movement protein of Pelargonium flower break virus shows RNA-binding activity and contains a biologically relevant leucine zipper-like motif

Martínez-Turiño, Sandra; Hernández, Carmen

doi:10.1016/j.virol.2011.03.001

Cited by 17 publications

(14 citation statements)

References 72 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…benthamiana and C . quinoa leaves were obtained by phenol extraction and LiCl precipitation [ 36 ]. In addition, for RNA stability assays, protoplasts were collected at 1 and 3 h after inoculation and total RNA was extracted using NucleoZOL reagent according the manufacturers instructions (Macherey-Nagel).…”

Section: Methodsmentioning

confidence: 99%

Efficient Translation of Pelargonium line pattern virus RNAs Relies on a TED-Like 3´-Translational Enhancer that Communicates with the Corresponding 5´-Region through a Long-Distance RNA-RNA Interaction

et al. 2016

Self Cite

View full text Add to dashboard Cite

Cap-independent translational enhancers (CITEs) have been identified at the 3´-terminal regions of distinct plant positive-strand RNA viruses belonging to families Tombusviridae and Luteoviridae. On the bases of their structural and/or functional requirements, at least six classes of CITEs have been defined whose distribution does not correlate with taxonomy. The so-called TED class has been relatively under-studied and its functionality only confirmed in the case of Satellite tobacco necrosis virus, a parasitic subviral agent. The 3´-untranslated region of the monopartite genome of Pelargonium line pattern virus (PLPV), the recommended type member of a tentative new genus (Pelarspovirus) in the family Tombusviridae, was predicted to contain a TED-like CITE. Similar CITEs can be anticipated in some other related viruses though none has been experimentally verified. Here, in the first place, we have performed a reassessment of the structure of the putative PLPV-TED through in silico predictions and in vitro SHAPE analysis with the full-length PLPV genome, which has indicated that the presumed TED element is larger than previously proposed. The extended conformation of the TED is strongly supported by the pattern of natural sequence variation, thus providing comparative structural evidence in support of the structural data obtained by in silico and in vitro approaches. Next, we have obtained experimental evidence demonstrating the in vivo activity of the PLPV-TED in the genomic (g) RNA, and also in the subgenomic (sg) RNA that the virus produces to express 3´-proximal genes. Besides other structural features, the results have highlighted the key role of long-distance kissing-loop interactions between the 3´-CITE and 5´-proximal hairpins for gRNA and sgRNA translation. Bioassays of CITE mutants have confirmed the importance of the identified 5´-3´ RNA communication for viral infectivity and, moreover, have underlined the strong evolutionary constraints that may operate on genome stretches with both regulatory and coding functions.

show abstract

Section: Methodsmentioning

confidence: 99%

Efficient Translation of Pelargonium line pattern virus RNAs Relies on a TED-Like 3´-Translational Enhancer that Communicates with the Corresponding 5´-Region through a Long-Distance RNA-RNA Interaction

et al. 2016

Self Cite

View full text Add to dashboard Cite

show abstract

“…In other words, the leucine zipper motif facilitates dimerization through parallel coiled-coil motifs and binds DNA through its hydrophobic domain. In more recent publications, this motif is no longer considered to be restricted to DNA binding but is generally accepted as a protein-protein interaction domain (24,32). To illustrate the domain in pUL71, we presented aa 34 to 55 in an alpha-helical wheel (data not shown).…”

Section: Fig 2 Oligomerization Potential Of Rpul71 (A)mentioning

confidence: 99%

A Leucine Zipper Motif of a Tegument Protein Triggers Final Envelopment of Human Cytomegalovirus

Meissner

Suffner

Schauflinger

et al. 2012

J Virol

View full text Add to dashboard Cite

bThe product of the human cytomegalovirus (HCMV) UL71 gene is conserved throughout the herpesvirus family. During HCMV infection, protein pUL71 is required for efficient virion egress and is involved in the final steps of secondary envelopment leading to infectious viral particles. We found strong indications for oligomerization of pUL71 under native conditions when recombinant pUL71 was negatively stained and analyzed by electron microscopy. Oligomerization of pUL71 during infection was further verified by native and reducing polyacrylamide gel electrophoresis (PAGE). By in silico analyses of the pUL71 sequence, we noticed a basic leucine zipper (bZIP)-like domain, which might serve as an oligomerization domain. We demonstrated the requirement of the bZIP-like domain for pUL71 oligomerization by coimmunoprecipitation and bimolecular fluorescence complementation using a panel of pUL71 mutants. These studies revealed that the mutation of two leucine residues is sufficient to abrogate oligomerization but that intracellular localization of pUL71 was unaffected. To investigate the relevance of the bZIP domain in the viral context, recombinant viruses carrying mutations identical to those in the panel of pUL71 mutants were generated. bZIP-defective viral mutants showed impaired viral growth, a small-plaque phenotype, and an ultrastructural phenotype similar to that of the previously described UL71 stop mutant virus. The majority of virus particles within the viral assembly compartment exhibited various stages of incomplete envelopment, which is consistent with the growth defect for the bZIP mutants. From these data we conclude that the bZIP-like domain is required for oligomerization of pUL71, which seems to be essential for correct envelopment of HCMV.

show abstract

“…Reverse genetics experiments have shown that both proteins are required for viral replication [24]. Two small ORFs, located in the central part of the viral genome, encode polypeptides of 7 (p7) and 12 kDa (p12), respectively, which are involved in virus movement [25] and the 3′-proximal ORF encodes a 37 kDa product (p37) which plays a dual role as coat protein (CP) and as suppressor of RNA silencing [26].…”

Section: Introductionmentioning

confidence: 99%

An Internal Ribosome Entry Site Directs Translation of the 3′-Gene from Pelargonium Flower Break Virus Genomic RNA: Implications for Infectivity

Fernández-Miragall

Hernández

2011

PLoS ONE

Self Cite

View full text Add to dashboard Cite

Pelargonium flower break virus (PFBV, genus Carmovirus) has a single-stranded positive-sense genomic RNA (gRNA) which contains five ORFs. The two 5′-proximal ORFs encode the replicases, two internal ORFs encode movement proteins, and the 3′-proximal ORF encodes a polypeptide (p37) which plays a dual role as capsid protein and as suppressor of RNA silencing. Like other members of family Tombusviridae, carmoviruses express ORFs that are not 5′-proximal from subgenomic RNAs. However, in one case, corresponding to Hisbiscus chlorotic ringspot virus, it has been reported that the 3′-proximal gene can be translated from the gRNA through an internal ribosome entry site (IRES). Here we show that PFBV also holds an IRES that mediates production of p37 from the gRNA, raising the question of whether this translation strategy may be conserved in the genus. The PFBV IRES was functional both in vitro and in vivo and either in the viral context or when inserted into synthetic bicistronic constructs. Through deletion and mutagenesis studies we have found that the IRES is contained within a 80 nt segment and have identified some structural traits that influence IRES function. Interestingly, mutations that diminish IRES activity strongly reduced the infectivity of the virus while the progress of the infection was favoured by mutations potentiating such activity. These results support the biological significance of the IRES-driven p37 translation and suggest that production of the silencing suppressor from the gRNA might allow the virus to early counteract the defence response of the host, thus facilitating pathogen multiplication and spread.

show abstract

A membrane-associated movement protein of Pelargonium flower break virus shows RNA-binding activity and contains a biologically relevant leucine zipper-like motif

Cited by 17 publications

References 72 publications

Efficient Translation of Pelargonium line pattern virus RNAs Relies on a TED-Like 3´-Translational Enhancer that Communicates with the Corresponding 5´-Region through a Long-Distance RNA-RNA Interaction

Efficient Translation of Pelargonium line pattern virus RNAs Relies on a TED-Like 3´-Translational Enhancer that Communicates with the Corresponding 5´-Region through a Long-Distance RNA-RNA Interaction

A Leucine Zipper Motif of a Tegument Protein Triggers Final Envelopment of Human Cytomegalovirus

An Internal Ribosome Entry Site Directs Translation of the 3′-Gene from Pelargonium Flower Break Virus Genomic RNA: Implications for Infectivity

Contact Info

Product

Resources

About